It has been shown that peripheral clearance of amyloid beta peptides reduces amyloid beta peptide levels in the brain through what has been termed a "sink effect". We have exploited this "sink effect" by showing that the peptidase neprilysin, when expressed peripherally on leukocytes reduced amyloid beta peptide levels in the brain of a hAPP transgenic mouse model of Alzheimer's disease. To obtain peripheral neprilysin expression we transplanted bone marrow stem cells transduced with lentivirus expressing neprilysin to a recipient hAPP transgenic mouse. We now propose to further develop peripheral expression of neprilysin as a therapeutic treatment for AD. Two specific aims directed at further developing peripheral neprilysin expression, without the need for bone marrow transplantation, are proposed: SPECIFIC AIM 1. To test the use of a single chain antibody-neprilysin (NEP) chimeric protein targeted to red blood cells to lower brain amyloid beta peptide levels. Chimeric proteins composed of a single chain antibody targeted to a mouse erythrocyte epitope fused with neprilysin will be used to produce erythrocyte bound neprilysin which will be tested for the ability to lower brain amyloid beta peptide levels. SPECIFIC AIM 2. To test the use of a myristoylated-neprilysin to attach NEP to hematopoietic cells. We will generate a form of neprilysin in which a myristoyl-peptide is linked to the C-terminus. This myristoyl-NEP will be used to produce peripheral NEP by insertion into the plasma membrane of hematopoietic cells and tested for the ability to lower brain amyloid beta peptide levels. We have as an overall objective the translation of one of these methods to the testing of human subjects. PUBLIC HEALTH RELEVENCE: The focus of this proposal is to develope a new therapeutic approach for treating Alzheimer's disease. We propose to express the enzyme neprilysin on blood cells where it can degrade plasma amyloid beta peptides leading to a lowering of brain amyloid beta peptides which are known to play a major role in causing Alzheimer's disease.